The Ron receptor tyrosine kinase has recently been shown to be overexpressed and activated in a cohort of human cancers, with the most compelling data found in breast cancer. Ron is overexpressed in approximately 50% of human breast cancers, and has been shown to be an independent predictor of both metastases and poor prognosis in women with this disease. While Ron overexpression appears to be an important factor in human breast cancer and metastasis, a significant gap exists in our knowledge about the signaling pathways that Ron activates in breast tumors, and about the importance of these pathways with respect to overall tumor growth and metastatic dissemination. Studies from our laboratory have shown that Ron overexpression selectively in the mammary epithelium leads to mammary tumorigenesis in female mice with 100% incidence and is associated with a high degree of breast cancer metastasis to the liver and lungs. In this model, mammary tumorigenesis is associated with elevated expression of Ron, increased receptor phosphorylation and increased Ron-associated tyrosine kinase activity. However, the degree of dependency of the Ron ligand, hepatocyte growth factor-like protein (HGFL), in this model remains unknown. Biochemical analyses of primary mammary tumors from these mice demonstrate that Ron overexpression is associated with elevated levels of tyrosine phosphorylated 2-catenin and with the upregulation of downstream 2-catenin target genes. Similar to Ron, 2-catenin expression is also associated with poor prognosis in breast cancer patients. Moreover, our preliminary data show an association between high Ron expression and high2-catenin levels in human breast cancer specimens. We also show that HGFL-induced Ron activation in human breast cancer cell lines induces rapid 2-catenin accumulation and tyrosine phosphorylation of2-catenin that is translocated to the nucleus. Based on our preliminary and published data we hypothesize that ligand-mediated activation of Ron augments 2-catenin signaling, leading to increased breast cancer growth, survival, and metastatic dissemination. To test this hypothesis three Specific Aims are proposed: (i) to delineate the role of the Ron ligand, HGFL, in breast cancer cell growth, survival and metastatic dissemination in vivo; (ii) to examine the in vivo biological significance of2-catenin expression in Ron-mediated mammary tumorigenesis and metastasis, and (iii) to determine the association, mechanism, and biological relevance of HGFL-induced, Ron-dependent 2-catenin tyrosine phosphorylation in breast cancer cells. In total, the studies outlined in this proposal will be the first to directly test the significance of ligand-induced Ron receptor activation during tumorigenesis as well as to define the molecular cross-talk between the Ron and 2-catenin signaling pathways in breast cancer. We will utilize cutting edge in vivo approaches involving the conditional loss of2-catenin, or a gene targeted loss of the Ron ligand, in the presence of Ron oncogene overexpression to analyze spontaneous breast tumor formation and metastasis in a clinically relevant murine model. PUBLIC HEALTH RELEVANCE: Excluding skin cancer, breast cancer is the most common cancer in women and is the second leading cause of cancer death in women. Approximately 180,000 new cases of invasive cancer will be diagnosed and about 40,000 women are estimated to die of this disease. As breast cancer incidence and death rates increase with age, the prevalence and morbidity of breast cancer are likely to increase with the aging population and the Veteran's Administration's expenditures and care for patients with this disease will subsequently increase accordingly. While significant effort and advancement into the treatment of this disease have occurred at the VA and other institutions, a significant gap still exists in our knowledge of new treatment options for patients with breast cancer. The focus of this proposal is to define the role of a novel protein, termed Ron, and Ron- dependent signaling pathways as potential new therapeutic targets to combat this disease.